Estimating reporting bias and validity of self-reports are a major issue in sexual behavior and condom use research. Incident STDs as biomarkers are expensive, and not useful in low-incident populations. We have developed a highly sensitive polymerase chain reaction (PCR) assay for semen Y-chromosome (Yc) sequences in vaginal fluid as a potential biomarker of sexual behavior and/or condom efficacy. The assay is highly sensitive and can be performed on a self-administered vaginal swab. Pilot data found that after unprotected intercourse followed by an abstinence period, Yc was detectable for 15 days, and the half-life was 3.4 days. In this proposal, we will further characterize the assay by enrolling 50 women in a clinical study to demonstrate "proof of concept", i.e. that after condom-protected intercourse no Yc is detectable in the post-coital vagina. 2). To define assay sensitivity under different conditions, we propose further research to evaluate characteristics of the vaginal microenvironment, which could affect Yc clearance. We hypothesize that menses and douching would shorten Yc clearance times. First we will evaluate the presence/absence of menses on Yc clearance by evaluating Yc clearance over a carefully monitored 2 week post-coital period in 120 women who will participate as their own control-they will participate twice (for a total of 224 cycles), comparing the effect of having their menstrual period during the measurement interval Yc clearance time compared to a time interval where menses did not occur. We will evaluate the effect of douching, a common practice in women in Baltimore at risk for STD, by comparing Yc clearance in 120 women after douching, compared to a repeated study interval in the same subject where they did not douche (a total of 224 study cycles). From our 1990-92 studies of condom use, we have archived cervicovaginal lavage fluid from 82 women reporting sex within 2 weeks and 100% condom use, and 322 who did not use condoms. These samples will be assayed for Yc and compared with demographic and behavioral data, and concomitant markers of cervical inflammation and STDs at time of visit. The outcomes will be compared to the findings from a previously published study that used STD incidence as the biomarker variable. Since Yc sampling requires only a vaginal swab, and is stable under field conditions, this test promises to be a practical and easily used biomarker for sexual behavior research.